The long term objectives of this proposal are to understand in detail the role of c-Myb in hematopoiesis and leukemogenesis. To achieve these goals, a thorough understanding of the molecular mechanisms for the regulation of c-myb expression in normal and leukemic cells is essential. The specific aims under this fellowship training are: (1) To clone the cDNA of CMAT, an inducible up-regulator for c-myb transcription during T-cell activation, and characterize its function in normal and leukemic T cells; (2) To determine whether a block to transcription elongation is involved in the regulation of c-myb expression in normal resting and activated T cells and in T-cell ALL cells. A yeast one-hybrid genetic system and other alternative strategies will be used to screen a cDNA library for the cDNA encoding the CMAT factor. The function of CMAT will be further characterized by co-transfection and in vitro DNA binding studies. The mRNA levels of CMAT in normal and in leukemic cells will be determined and correlated with the levels of c-myb expression. The specific roles of CMAT and/or other factors in T-cell ALL cells will be determined by in vivo footprinting, gel mobility shift and UV-cross link studies and by comparing the results with hose found in normal cells and in cell lines. S1 nuclease mapping of the 3' ends of the nuclear c-myb RNAs will be used to investigate whether premature termination of c-myb transcription occur. Kinetic nuclear run-on and potassium permanganate in vivo footprinting techniques will be used to locate the inv vivo pause site of RNA polymerase II during the transcriptional elongation of c-myb.